This study aimed to formulate a milk protein concentrate (MPC) and evaluate the effect of this MPC supplementation on body composition, lipid profile, and product acceptability in resistance training (RT) practitioners. The physical, chemical and sensory properties were then analyzed. To evaluate the effect of the MPC supplementation on resistance training practitioners, 30 males were divided into two groups: a control group and an intervention group. Then the participants in the intervention group were supplemented with 500 mL of the MPC (250 mL pre-RT and 250 mL post-RT). Body composition (weight, height, and skinfolds) and biochemical markers (cholesterol and triglycerides) were measured at baseline and at 30, 60, and 90 days after treatment. Body composition results showed that the MPC supplementation during a RT program significantly (p < 0.05) reduced body fat percentage and fat mass, and increased lean mass in RT practitioners. Characterization of the MPC revealed low fat (3.32 ± 0.08%) and carbohydrates (4.58 ± 0.59%), and high protein (18.51 ± 0.51%) concentrations. Additionally, the product showed satisfactory acceptability index (> 71%). Thus, the MPC supplementation contributed to reduction of fat percentage, fat mass, and increase of lean mass in RT practitioners, without lipid profile alterations.